Thioredoxin is overexpressed in half of human primary lung, colon, and gastric cancers, and in a third of adult acute lymphocytic leukemias (ALL). The studies outlined in this proposal examine more closely the role of thioredoxin in human leukemic cell apoptosis. The hypothesis being tested is that thioredoxin is a new anti-apoptosis gene whose overexpression is associated with aggressive tumor growth and resistance to apoptosis. Our work with WEHI7.2 cells, a mouse lymphoid cell line, will be extended by developing cells that express a dominant negative redox inactive thioredoxin to examine more rigorously the in vitro and in vivo effects of thioredoxin expression on apoptosis and tumor formation. The most likely mechanism through which thioredoxin overexpression exerts its anti-apoptotic effect is via NF-kappaB activation. This will be tested by the development of WEHI7.2 cell lines that overexpress both thioredoxin and a dominant negative mutant of IkappaBalpha, which antagonizes NF-kappaB activation. Lastly, the subcellular distribution of thioredoxin may also play a role in the anti-apoptotic effect of thioredoxin overexpression. This will be tested by targeting thioredoxin overexpression specifically to the mitochondria, where the initial steps of the apoptotic program occur. The key to understanding the origins of human cancer is a detailed knowledge of the signaling pathways that mediate apoptosis and how they become altered in cancer. Such knowledge could also provide the basis for novel strategies to prevent and treat cancer and for overcoming drug resistance.